Visualization of Microscopy‐Based Spectral Imaging Data from Multi‐Label Tissue Sections

Combining images taken with light of specific wavelengths can dramatically enhance light‐microscopic images. This technology is enabled by the availability of programmable filters that can be set to transmit light only of particular wavelengths. Spectral imaging technologies have become an important part of microscopy, and are particularly useful for analyzing samples that have been labeled with multiple (two or more) molecular markers. The most commonly used methodology for separating the markers from each other is linear unmixing, which results in a quantitative image of the location and amount of each marker present in the sample. The very complexity of these multilabel samples requires a high degree of sophistication in methods to visualize the results of unmixing. This article describes a wide range of useful visualization tools designed to better enable discrimination of different features in multilabeled tissue or cell samples. These commercially available tools can be attached to the standard laboratory light microscope to significantly enhance the power of light microscopy. Curr. Protoc. Mol. Biol. 84:14.19.1‐14.19.15. © 2008 by John Wiley & Sons, Inc.